Deoxycholic acid (DCA) has been reported to play a role in the etiology of colon cancer and cholesterol gallstone disease in man. High levels of hydrophobic secondary bile acids may also contribute to the pathophysiology of certain cholestatic liver diseases. DCA is formed from cholic acid (CA) by a small population of gram-positive intestinal anaerobic bacteria. The levels of DCA in human bile can range from 0 to >40%. CA conversion to DCA occurs via a six step biochemical pathway. Th genes encoding enzymes in the bile acid 7alpha-dehydroxylation (7alpha-DeOH) pathway are located on a large bile acid inducible (bai) operon in Eubacteriim sp. VPI 12708. However, preliminary data indicates that the bai genes hybridize to DNA from only about 50% of other 6alpha-DeOH intestinal bacteria. Specific Aim1 is to clone, sequence and analyze the bai operon from Clostridum sp. Strain T0931, a high activity strain whose DNA does not hybridize to bai genes from Eubacterum sp. VPI12708 . This information will be essential for a more complete understanding of the origin and evolution of enzymes involved in bile acid 7alpha- DeOH. Specific Aim 2 is to express E. Coli, purify and characterize the baiF gene product for Eubacterium sp. VPI 12708. Preliminary data indicate that this gene encodes a novel bile acid-CoA hydrolase. A better understanding of the genetics and enzymology of bile acid 7alpha-DeOH is essential for the development of specific inhibitors of this pathway.